Abstract

The microbiome affects development and activity of the immune system, and may modulate immune therapies, but there is little direct information about this control in vivo. We studied how the microbiome affects regulation of human immune cells in humanized mice. When humanized mice were treated with a cocktail of 4 antibiotics, there was an increase in the frequency of effector T cells in the gut wall, circulating levels of IFN-γ, and appearance of anti-nuclear antibodies. Teplizumab, a non–FcR-binding anti-CD3ε antibody, no longer delayed xenograft rejection. An increase in CD8+ central memory cells and IL-10, markers of efficacy of teplizumab, were not induced. IL-10 levels were only decreased when the mice were treated with all 4 but not individual antibiotics. Antibiotic treatment affected CD11b+CD11c+ cells, which produced less IL-10 and IL-27, and showed increased expression of CD86 and activation of T cells when cocultured with T cells and teplizumab. Soluble products in the pellets appeared to be responsible for the reduced IL-27 expression in DCs. Similar changes in IL-10 induction were seen when human peripheral blood mononuclear cells were cultured with human stool samples. We conclude that changes in the microbiome may impact the efficacy of immunosuppressive medications by altering immune regulatory pathways.

Figure 8

(A–C) Splenocytes from mice that had not been treated with antibiotics (Abx) were cultured with pools of pellets from mice that had not or had been treated with antibiotics. The levels of IFN-γ, IL-10, and TNF were measured in the supernatants after 5 days and compared by paired t test. *P = 0.04, n = 8 cell donors/group. (D) Subsets of human PBMCs were isolated by magnetic beads and cultured with pellets from mice that had or had not been treated with antibiotics (n = 5 each). There was increased IL-10 release when the DCs were cultured with pellets from both groups of mice but reduced release with pellets from the antibiotic-treated mice. *P = 0.03, ***P = 0.0008, ****P < 0.0001, 2-way ANOVA with paired comparisons. (E–I) DCs were isolated from the PBMCs from healthy donors and cytokine levels were measured after 5 days in culture with a pool of pellets from mice that had or had not been treated with antibiotics. Each symbol represents a DC cell donor. Culture with the pellets increased production of all cytokines (P < 0.0001) compared with the cultures without pellets (Ctrl) (P = 0.002 by ANOVA). (J) Human stool samples of antibiotic-treated patients and healthy controls were heat inactivated and cultured with PBMCs that were pooled from 4 healthy cell donors. The level of IL-10 in the supernatant after culture for 3 days is shown. Each symbol represents a stool sample donor. **P = 0.002, ****P < 0.0001 by Student’s t test.